The risk of sexually transmitting a chronic hepatitis C infection to a long-term monogamous heterosexual partner is very low, averaging just about 1% per year.

That risk level works out to a transmission rate of about one in every 190,000 sexual contacts, Dr. Norah Terrault and her colleagues reported in the April issue of Hepatology (2013;57:881-9).

The cross-sectional study also found that no one sexual practice – including anal intercourse or intercourse during menses – significantly increased the risk of transmission, wrote Dr. Terrault of the University of California, San Francisco. The findings can be used to provide "unambiguous and reassuring counseling messages," she and her coinvestigators noted.

Each of the partners was interviewed separately about their sexual contacts and practices. At the time of interview, the index subjects were a median of 49 years old and the partners, a median of 48 years.

The HCV-positive subjects reported the highest incidence of past risk factors, including blood transfusions before 1992 (32%), injected illegal drugs (54%), and being stuck by a bloody sharp item in a hospital (4%). Nearly half (46%) reported having had at least 20 lifetime sexual partners, with 21% having had 50 or more.

However, partners also reported some risk factors: 11% had an early transfusion, 2% used illegal drugs, and 2% had a hospital sharps incident. Many (27%) also reported having had at least 20 sexual partners.

Among the 500 couples, 20 partners (4%) were coinfected with HCV. Of these, nine were concordantly infected, eight discordantly, and three were indeterminate.

Six of the concordant couples underwent phylogenetic typing. Three were infected with the same HCV isolate and three with different strains. The investigators estimated the time of transmission and any additional risk factor among the three couples with concordant strains.

For the first couple, with an 18-year relationship, transmission probably occurred after about 6.5 years. The female partner had a history of injected drug use, while the male had no identifiable risk factors.

The second couple had a 28-year relationship; transmission probably occurred at around 15 years, the investigators said. "The female partner had a history of injectable drug use and both partners reported more than 20 prior sexual partners, a history of sexual transmitted diseases, and a history of snorting of drugs."

For the third couple, who had been together for 10 years, transmission probably occurred at around year 6. "The male partner had a history of injectable drug use, of being stuck by a sharp bloody object while working in a hospital, and more than 20 prior sexual partners; both partners reported snorting drugs and sharing snorting equipment."

The investigators determined that these infections were probably sexually transmitted between the partners – a prevalence of about 1%. "The estimated risk per sexual contact ranged from 1/380,000 to 1/190, 000," they said.

However, they were unable to identify any behaviors that significantly increased the risk of transmission. Compared with couples without coinfection, coinfected couples were more likely to have vaginal intercourse during menses (100% vs. 66%), more likely to have anal intercourse (67% vs. 30%), and less likely to use condoms (0% vs. 30%), but none of these differences was statistically significant.

"HCV transmission by sex from chronically infected persons to their heterosexual partners in a long-term monogamous relationship likely occurs, but is a rare event," the authors concluded. "Our results provide a basis for specific counseling messages that clinicians can use with their patients... [that] support the current national recommendations that couples not change their sexual practices if they are in a monogamous heterosexual relationship."

Governments across the world will have to face up to the challenges posed by the hepatitis C (HCV) pandemic or experience spiralling healthcare costs, says the Economist Intelligence Unit in its report The silent pandemic: Tackling hepatitis C with policy innovation.

The report, made possible as a result of an educational grant from Janssen, challenges countries to use co-ordinated strategies to tackle HCV but warns that this will not be easy because few countries understand the magnitude of the disease. The report finds that in the European Union, only the Netherlands has the kind of epidemiological data robust enough to inform policy.

What the report highlights is that because levels of awareness are so low, many people only receive an HCV diagnosis when diagnosed with its end-stage conditions such as cirrhosis or liver cancer. Poor healthcare practices, such as failing to screen donated blood and the use of unsterilised medical equipment, are the cause of millions of cases in the developing world.

The report concludes that countries need to improve the data they have in order to introduce a comprehensive approach to tackling HCV. This will include raising awareness about the disease, taking preventative measures, especially within health services, and making a real effort to reach vulnerable patient groups with available treatments treatments before end-stage conditions develop.

International Coalition of Hepatitis C Survivors band together to raise awareness, fund research, and find a cure

NEW YORK, June 17, 2013 /PRNewswire via COMTEX/ -- Today in New York the World Hepatitis Fund launched a bold new initiative to harness the power of social media to educate, diagnose and treat the nearly two hundred million people worldwide who have contracted Hepatitis C.

Hepatitis C, also known as the silent killer is the leading cause of liver cancer and the need for liver transplantation. Transmitted via blood, it often goes undetected for years, even decades, without symptoms. In the United States, where an estimated three million people are infected with Hepatitis C, the Centers for Disease Control (CDC) recommends that every adult born from 1942 to 1965 be screened for the HCV virus in order to lead them to treatment.

The World Hepatitis Fund is the brainchild of Humberto Silva, who himself was diagnosed with the disease by a routine screening, "I didn't know I was infected, and my four children have a father because of a simple blood test," said Silva, "it is my mission to get everyone educated, tested and treated."

Silva is joined on his mission by an international coalition of Hepatitis C survivors; they currently fund the project and are working to develop international guidelines. The Fund will be based in the United States, with offices in New York City, satellite offices in Brazil, Egypt, Pakistan, Colombia, Japan, China and England.

For more information, and to get involved visit the World Hepatitis Fund on Facebook.

Received 26 December 2012; received in revised form 6 February 2013; accepted 12 February 2013. published online 25 February 2013.

Abstract

Background & Aims

ABT-450 (combined with low-dose ritonavir, ABT-450/r) is a potent HCV NS3 protease inhibitor, and ABT-072 is a non-nucleoside NS5B polymerase inhibitor. The goal of this study was to evaluate the safety, tolerability, and efficacy of the peginterferon-free combination of ABT-450/r and ABT-072 with ribavirin in treatment-naïve patients with IL28B CC genotype, infected with HCV genotype 1.

Conclusions

A 12-week regimen of ABT-450/r and ABT-072 with ribavirin was well tolerated with 9/11 patients achieving sustained virologic response through 36weeks of post-treatment observation. These findings suggest that peginterferon-free regimens may have the potential to cure a high proportion of HCV genotype 1-infected patients.

Received 26 November 2012; received in revised form 4 February 2013; accepted 12 February 2013. published online 22 February 2013.

Abstract

Background & Aims

MK-7009 (vaniprevir) is a non-covalent competitive inhibitor of the hepatitis C virus (HCV) NS3/4A protease. This report presents the primary analysis results (safety and sustained viral response) of a phase 2b study of MK-7009 given in combination with peginterferon (PegIFN) alfa2a 180μg weekly and ribavirin (RBV) 1000–1200mg/day, for 24–48weeks to non-cirrhotic patients who have failed previous PegIFN and RBV treatment.

Methods

We present results of a randomized, placebo-controlled, double-blind study of MK-7009 administered for 24–48weeks in combination with PegIFN and RBV in 4 regimens to at least 40 patients per arm. Stratification by prior response to PegIFN and RBV was as follows: null response, partial response, breakthrough and relapse. HCV RNA was determined by Roche Cobas Taqman with a lower limit of detection (LLoD) of 10IU/ml and a lower limit of quantification (LLoQ) of 25IU/ml.

Results

SVR24 in patients on MK-7009+PegIFN and ribavirin (P/R) was statistically superior to placebo+P/R in all treatment groups (p<0.001). MK-7009 at 300mg b.i.d. and 600mg b.i.d. is generally well tolerated for use for up to 48weeks of therapy. Patients in MK-7009 regimens had higher rates of gastrointestinal adverse events as compared to control (mostly mild to moderate). There were no significant differences in rates of anemia and rash between the MK-7009 regimens and control.

Conclusions

In conclusion, patients treated with MK-7009 plus P/R experienced significant improvement in SVR compared to P/R control in a population of GT 1 experienced patients.

Patients coinfected with chronic hepatitis C and occult hepatitis B were more likely to develop cirrhosis or hepatocellular carcinoma and had poorer survival than patients with hepatitis C alone in a recent study.

In an observational cohort study, researchers tested 326 patients with chronic HCV for occult HBV infection (OBI) between 1991 and 2000. All participants were hepatitis B surface antigen (HBsAg)-negative and underwent liver biopsy. Follow-up for a median of 11 years (range 5-19 years) was performed in 94 patients, including 37 OBI-positive and 57 OBI-negative participants.

Seventy-nine participants received interferon-based HCV therapy, either with or without ribavirin. Sustained virologic response occurred in 26 patients, independently of OBI status.

Hepatocellular carcinoma (HCC) developed in 13 OBI-positive and five OBI-negative participants across a median of 8.8 years (P<.01). In this group, patients with OBI were younger than those without OBI (60 years vs. 74 years; P<.05). Among participants who did not develop HCC, eight OBI-positive and seven OBI-negative participants developed advanced cirrhosis. Worsening liver disease was directly correlated with OBI via the Spearman Correlation Test (P<.001).

Across the entire cohort, 18 participants died and two underwent liver transplantation. All deaths were due to liver-related causes. Cumulative survival rates were shorter (P=.003) and liver-related deaths were more frequent among those with OBI (12 cases vs. six; P<.01). Investigators noted associations between poor survival and HCC development (P<.01) and lack of response to HCV therapy (P=.02).

“The observation that response to anti-HCV therapy (that was not influenced by the OBI status) is associated with a benign evolution of the liver disease, thus possibly nullifying the negative effect of OBI on the liver disease outcome, is of the utmost importance,” the researchers wrote. “Altogether, these data and considerations may lead to the conclusion that, among [chronic HCV] patients, the occult HBV coinfected individuals represent a category at high risk of progression toward cirrhosis, HCC development and lower survival, thus representing a subset of patients in whom curing the HCV infection appears to be a high priority.”

In patients with chronic Hepatitis C virus (HCV) infection, a sustained viral response to treatment regimens 12 weeks after therapy (SVR12) is a good indicator that the response will be maintained until week 24 (SVR 24), based on an analysis of pooled clinical trial data published in the June issue of Gastroenterology. Therefore SVR12 can be used instead of SVR24 as a primary end point for registration trials.

A SVR24 (undetectable levels of HCV RNA in serum 24 weeks after completion of therapy) is considered to indicate successful treatment—most patients who achieve SVR24 maintain their serum-negative status and have reduced complications from liver disease and increased survival times. It is therefore the primary endpoint of HCV therapy trials for regulatory approval—efficacies of all HCV therapies approved by the US Food and Drug Administration have been based on the proportion of patients attaining SVR24. However, earlier time points could increase efficiency of drug development.

Jianmeng Chen et al. assessed data from 15 phase 2 and 3 trials, 3 pediatric studies, and 5 drug-development programs to determine the concordance between SVR24 and SVR12, and even SVR4. They analyzed data from groups of subjects who received various combinations and regimens with interferon, ribavirin, and direct-acting antiviral agents.

Of the 13,599 adult subjects in the database analyzed, 50.6% achieved SVR24 and 51.8% achieved SVR12. These appeared to be mostly the same patients—the positive predictive value of SVR12 for SVR24 was 98% and the negative predictive value was 99% among subjects with genotype 1 HCV infection. A similar level of concordance was observed in subjects with HCV genotype 2 or 3 infections, as well as in pediatric studies.

Why is there such a high level of agreement between SVR12 and SVR24, and to a lesser extent SVR4 and SVR24? Chen et al. explained that 65% of subjects who relapsed had detectable viral load by week 4 after treatment and 95% had detectable viral load by week 12. So if the virus is going to reappear, in generally does so within 12 weeks after the treatment ends.

The authors conclude that individual patients should be followed for 24 weeks or longer after treatment to confirm their responses. But SVR12, which is currently used as supportive information for registration trial design, can also be appropriate for regulatory approval of treatment regimens.

Current figures for Thailand put the number of infected individuals at more than one million but most experts estimate that this is on the low side, as the disease is asymptomatic, so those infected are only likely to consult a doctor if they fall ill with another condition.

Research has also snown that the virus appears to have a geographical bias with both the north and northeast reporting a three to six per cent infection rate compared to just one to two per cent in the central region.

Hepatitis C causes chronic liver inflammation and continually destroys the liver cells, causing liver fibrosis and eventually leading to cirrhosis. This causes the liver to function abnormally, an increase in the AST and ALT levels and a gradual deterioration in the liver function, leading to chronic liver failure. While patients will have no symptoms until late in the disease, at this stage they will develop signs of jaundice, yellow eyes, ascites (build-up of fluid in the abdominal cavity) and neurological conditions as result of kidney failure. The virus can also cause liver cancer especially among patients already suffering from cirrhosis.

About 70 to 85 per cent of Hepatitis C cases can be treated through a combination of pegylated interferon injections, which are administered once a week along with oral Ribavirin.

Duration and success of the medication depends on the strain of the hepatitis C virus. Patients infected with strains 2 and 3 of the virus require 24 weeks of treatment and the chances of being cured are 80 to 85 per cent. Those infected with virus strain 1 require 48 weeks of treatment and the chance of being cured stands at 70 per cent. Patients infected with virus strain 6 require 24-48 weeks of treatment and the cure rate is 70 to 76 per cent

Two anti-retrovirals, Boceprevir and Telaprevir, were approved by the USFDA in 2011 as oral medications for strain 1 of the virus and have proved particularly useful in increasing the efficiency of the pegylated interferon injection with Ribavirin, especially in patients who have not responded well to the current treatment option. Research is being undertaken on several other anti-retroviral drugs that can used for other strains of the virus and it is hoped that before long, treatment will involve only oral anti-retroviral medications rather than the combination of oral and injected.

This will make the treatment more convenient, create fewer side effects, avoid the use of injections and increase the chances of recovery.

But to treat patients, first they must be diagnosed and for that reason, the Liver Foundation and Liver Society of Thailand are organising a campaign to educate the public about Hepatitis C and its dangers.

Dr Teerha Piratvisuth is a specialist in liver and gastroenterology at Samitivej Sukhumvit Hospital. Call the Liver and Digestive Institute at (02) 711-8822-4 or visit "samitivej" on Facebook.

Vocal New York, a grassroots advocacy group representing low income residents, and Assemblyman Kenneth Zebrowski, are pushing for the passage of Hepatitis-C testing legislation in the Senate, with only a few days left in session. Photo by Cassandra Hamdan.

People in black and white tee-shirts with Vocal New York printed across their fronts crowded the Million Dollar Staircase in the Capitol; signs, that said, "Hep C Testing = Saved Lives," and "Fight Hep C," waved in the air; chants of, "No justice, no peace," and "End Hep-C" reverberated up and down the staircase; Tuesday was a day to make history in the eyes of some 50 people.

"You are all making history here," Assemblyman Kenneth Zebrowski, D-New City, told the group of advocates from Vocal New York, a grassroots advocacy group that speaks on the behalf of low-income people, and concerned individuals as they rallied behind a bill (A.1286/S.2750) — the first piece of legislation of its kind nationally — to add a new section to the public health law requiring certain health service providers to offer Hepatitis-C tests to people born between 1945 and 1965. The baby boomer generation has the highest prevalence of Hepatitis-C of any other generation, with 1 out of 30 having Hepatitis-C, according to the Centers for Disease Control and Prevention. Zebrowski said that, though the CDC is not 100 percent sure why baby boomers are so much more susceptible, he believes it's because blood was not typically tested before widespread transfusions began in the early 1990s

Under the bill, if the test should come out positive, the insurance provider must offer follow-up health care or refer the infected individual to a provider who can, including a Hepatitis-C diagnostic test.

The bill would also require the state health commissioner to evaluate the impact of the legislation and report the findings to the governor and the Legislature.

The bill passed in the Assembly on June 10 and advanced into its third reading in the Senate on the same day. The bill was sponsored in the Assembly by Zebrowski whose father, also an assemblyman, died in 2007 of Hepatitis-C. "Three to five million people have Hep-C and most of them don't even know," Zebrowski said. He seemed optimistic about the possibility of the bill passing in the Senate, saying that all it needs is a "little push."

"We will get this all signed into law."

Assemblywoman Joan Millman, D-Brooklyn, who co-sponsors the bill, said her message was simple, "We need to pass this bill."

The rally was emceed by Bobby Tolbert, a board member of Vocal New York, who opened the floor to personal testimonies. One such testimony was given by a Diane Nunez who was diagnosed with Hepatitis-C in 1998 and went for treatment in 2003. "This is a pandemic in our communities," said Nunez, "We have to end Hep-C."

Hadiyah Charles, a longtime HIV/AIDS prevention advocate associated with the Lower East Side Harm Reduction Coalition, praised state lawmakers for taking a big step in national history and said that with this legislation "New York should cause a domino effect" of other states passing similar legislation.

Sammy Santiago, who was tested in 1996 for Hepatitis-C and was declared "undetectable," earlier this year found out he had cirrhosis of the liver. He has been getting treatment for Hepatitis-C for four months. "In the first four weeks I was declared undetectable again," Santiago said. Santiago stressed the importance of getting tested for Hepatitis-C, "That's what's important; we all need to be educated."

Assemblyman Zebrowski said he remains optimistic that the Senate will pass the bill.

George Santana of the CitiWide Harm Reduction Coalition spoke about his experience after being diagnosed with Hepatitis-C. "I've done the treatment. It sucks, anyone who has done it knows what I'm saying, but it was worth it," Santana said. "We must continue to fight, because I know deep in my heart this bill will pass."

As the first major clinical reference laboratory to offer the COBAS AmpliPrep/COBAS TaqMan HCV Test, v2.0, LabCorp can now provide physicians with a quantitative HCV viral load assay that has a lower limit of detection and quantification (15 IU/mL) than existing qualitative HCV assays (10-50 IU/mL). Viral load determinations influence decisions related to many aspects of antiviral HCV therapy, including treatment selection and adjustments. Following treatment initiation, periodic measurements of HCV viral load allows the clinician to assess the success of treatment.

An estimated 3.2 million people in the U.S. (and 170 million worldwide) are chronically infected with HCV, which if left undiagnosed and untreated can lead to liver fibrosis, cirrhosis and hepatocellular carcinoma. The Centers for Disease Control and Prevention (CDC) estimates that nearly half of the U.S. HCV population is currently undiagnosed, and the slow and often silent onset of HCV disease presentation has prompted more aggressive efforts to proactively diagnose and treat HCV infection. Recently, the CDC expanded its HCV screening recommendations from “high risk behavior groups” to include all individuals born between 1945-1965, a birth cohort that CDC estimates to include a majority of HCV infected individuals in the U.S. As part of this initiative, newly diagnosed individuals are encouraged to seek medical care and treatment for their HCV infection. The expanded availability through LabCorp of the COBAS AmpliPrep/COBAS TaqMan HCV Test, v2.0 is particularly timely, as the CDC recommendation is expected to result in the identification of additional HCV-infected patients and an increased demand for antiviral treatment regimens.

“LabCorp is proud of its longstanding and prominent role of providing innovative and novel diagnostic tests to assist physicians in treating and managing chronic HCV infection,” said Dr. Mark Brecher, LabCorp’s Chief Medical Officer. “The COBAS AmpliPrep/COBAS TaqMan HCV Test, v2.0 is another valuable addition to the Company’s comprehensive portfolio of assays that characterize the Hepatitis C virus, disease course and the patient’s optimal treatment path.”

This press release contains forward-looking statements. Each of the forward-looking statements is subject to change based on various important factors, including without limitation, competitive actions in the marketplace and adverse actions of governmental and other third-party payors.Actual results could differ materially from those suggested by these forward-looking statements. Further information on potential factors that could affect LabCorp’s financial results is included in the Company’s Form 10-K for the year ended December 31, 2012, and subsequent SEC filings.

Age disparities exist in the continuum of care for patients with the human immunodeficiency virus (HIV) with people younger than 45 years less likely to be aware of their infection or to have a suppressed viral load, according to a report published Online First by JAMA Internal Medicine, a JAMA Network publication.

Early diagnosis, prompt and sustained care, and antiretroviral therapy (ART) are associated with reduced morbidity, mortality and further transmission of the virus. However, of the more than 1.1 million people living with HIV, more than 200,000 are unaware they are infected, less than 50 percent of people infected receive regular care and fewer than 30 percent have a suppressed viral load, the authors write in the study background.

H. Irene Hall, Ph.D., of the Centers for Disease Control and Prevention, Atlanta, and colleagues used data from the National HIV Surveillance System to determine the number of people living with HIV who are aware and unaware of their infection. Researchers also calculated the percentage of people linked to care within three months of diagnosis and estimated the percentages of people who were retained in care and who were prescribed ART.

"Additional efforts are needed to ensure that all persons with HIV receive a diagnosis and optimal care to reduce morbidity, mortality, disparities in care and treatment, and ultimately HIV transmission," the authors note.

Of the estimated more than 1.1 million persons living with HIV in 2009, nearly 81.9 percent had been diagnosed, 65.8 percent were linked to care and 36.7 percent were retained in care, 32.7 percent were prescribed ART and 25.3 percent had a suppressed viral load, according to study results.

Among people infected with HIV who were 13 to 24 years of age, 40.5 percent had received a diagnosis and 30.6 percent were linked to care. Lower percentages of people ages 25 to 44 were retained in care, were prescribed ART and had a suppressed viral load than were people ages 55 to 64 years of age. For example, among patients ages 25 to 34 years, 28 percent were in care compared with 46 percent among those patients ages 55 to 64 years, the results indicate.

Overall, 857,276 patients with HIV had not achieved viral suppression, including 74.8 percent of male, 79 percent of black, 73.9 percent of Hispanic/Latino and 70.3 percent of white patients, the results also show.

"Individuals, health care providers, health departments and government agencies must all work together to increase the numbers of people living with HIV who are aware of their status, linked to and retained in care, receiving treatment and adherent to treatment," the authors conclude.

(JAMA Intern Med. Published online June 17, 2013. doi:10.1001/jamainternmed.2013.6841. Available pre-embargo to the media at http://media.jamanetwork.com.)

Editor's Note: The CDC provides funds to all states and the District of Columbia to conduct the HIV surveillance used in this study. Please see the article for additional information, including other authors, author contributions and affiliations, financial disclosures, funding and support, etc.

Commentary: Overcoming the HIV Obstacle Course

In an invited commentary, Katerina A. Christopoulos, M.D., M.P.H., and Diane V. Havlir, M.D., of the University of California, San Francisco, write: "In 2011, the HIV field was shocked to learn that only about a quarter of individuals living with HIV were successfully receiving HIV treatment."

"The sobering numbers of those missing out on effective treatment because they did not know they were infected and those who knew their status but did not seek care spurred collaboration between the HIV treatment and prevention movements, two areas with different funding streams that often operated independently of one another," they continue.

"Already the HIV community has mobilized to further develop and study interventions that address bottlenecks in the cascade. Achieving an AIDS-free generation will be within reach if, and only if, these efforts succeed," they conclude.

(JAMA Intern Med. Published online June 17, 2013. doi:10.1001/jamainternmed.2013.7943. Available pre-embargo to the media at http://media.jamanetwork.com.)

Editor's Note: One of the authors made a conflict of interest disclosure. Please see the article for additional information, including other authors, author contributions and affiliations, financial disclosures, funding and support, etc.

Abstract and Introduction

Abstract

Study C209 evaluated the activity of telaprevir in treatment-naïve patients with genotypes 2 or 3 (G2, G3) hepatitis C virus (HCV) infection. Telaprevir monotherapy showed potent activity against HCV G2, but limited activity against G3. This analysis was performed to characterize HCV viral variants emerging during telaprevir-based treatment of G2/G3 HCV-infected patients. Patients were randomized to receive 2 weeks of treatment with telaprevir (telaprevir monotherapy), telaprevir plus peginterferon alfa-2a and ribavirin (triple therapy), or placebo plus peginterferon alfa-2a and ribavirin (control), followed by 22–24 weeks of peginterferon/ribavirin alone. Viral breakthrough was defined as an increase >1 log10 in HCV RNA from nadir, or HCV RNA >100 IU/mL in patients previously reaching <25 IU/mL. Twenty-three patients (47%) had G2 and 26 (53%) had G3 HCV. Viral breakthrough occurred during the initial 2-week treatment phase in six G2 patients (66.7%; subtypes 2, 2a and 2b) and three G3 patients (37.5%; all subtype 3a), all in the telaprevir monotherapy arm. Four breakthrough patients (three G2, one G3) subsequently achieved sustained virologic response (SVR). In all patients with breakthrough and available sequence data, mutations associated with reduced susceptibility to telaprevir in genotype 1 (G1) HCV were observed. No novel G2/G3-specific mutations were associated with telaprevir resistance. The telaprevir resistance profile appeared consistent across HCV genotypes 1, 2 and 3. Although viral breakthrough with resistance occurred in patients receiving telaprevir monotherapy, half of these patients achieved an SVR upon addition of peginterferon/ribavirin highlighting the importance of combination therapy.

Introduction

Chronic hepatitis C virus (HCV) infection is a major global healthcare burden.[1] Six HCV genotypes and over 100 subtypes are documented,[2] each with a distinct geographical distribution. Genotype 1 (G1) HCV predominates in Europe, the United States and Japan, G2 predominates in Mediterranean countries and the Far East, and G3 is common in the Indian subcontinent.[3,4] An increase in the proportion of patients with G3 HCV infection was recently observed in Europe.[5]

Two direct-acting antiviral (DAA) agents that target the HCV NS3-4A serine protease, telaprevir[6] and boceprevir,[7] showed substantial efficacy against chronic HCV G1 infection[8–12] and were recently approved in the United States and Europe. Preliminary in vitro data suggested that telaprevir also has activity against G2/G3 HCV.[13] Therefore, the Phase IIa C209 study evaluated the activity of telaprevir, administered for 15 days with or without peginterferon/ribavirin, on G2/G3 HCV infection. When administered as monotherapy, telaprevir had significant and rapid antiviral activity against G2 HCV, but little or no activity against G3.[14]

Primary and secondary endpoints are reported elsewhere.[14] Viral breakthrough (vBT) was defined as an increase >1 log10 in HCV RNA from nadir or HCV RNA >100 IU/mL in patients who previously reached HCV RNA <25 IU/mL, confirmed by two consecutive samples. Patients who had an unconfirmed vBT during telaprevir monotherapy were still considered to have vBT. SVR was defined as undetectable HCV RNA at end of treatment (EOT) and 24 weeks afterwards.

The study was approved by each centre's institutional review board and conducted in accordance with the Declaration of Helsinki and Good Clinical Practice guidelines. Written informed consent was obtained from all patients.

Samples for viral sequencing were obtained at baseline; days 2, 3, 4, 8, 12 and 15; weeks 4, 6 and 14; at EOT; 4, 8, 12 and 24 weeks after EOT; and 24 weeks after relapse (if any). Population-based NS3 sequence analysis was conducted in all baseline samples and in postbaseline samples from telaprevir-treated patients with vBT and/or not achieving an SVR. Population-based sequencing of the HCV NS3 region used subtype-specific amplification and sequencing protocols (Virco BVBA, Beerse, Belgium). Clonal sequencing of the NS3 region was conducted in a subset of samples. For both population-based and clonal sequencing, total RNA was extracted from plasma. The RNA was reverse transcribed, and cDNA was amplified in a one-step polymerase chain reaction (PCR) using the SuperScriptTM III One-Step Reverse Transcriptase (RT)-PCR System (Life Technologies, Carlsbad, CA, USA). An amplicon encompassing the NS3 region was further amplified during a nested PCR (Expand high Fidelity PCR system, Roche Applied Science, Penzberg, Germany). For population-based sequencing, PCR products were purified and directly sequenced in both directions using Big Dye terminator sequencing chemistry (Life Technologies). In case of clonal sequencing, the PCR product was first cloned using a TOPO TA cloning kit (Life Technologies). DNA from 20 to 30 clones per sample was purified and sanger sequenced as described above. The genotype and subtype of each sample were determined based on the NS5B region. The lower limit of detection for the sequencing assay was ~1000 IU/mL HCV RNA.

Statistical Analyses

To identify mutations potentially associated with reduced susceptibility to telaprevir, the frequency of every amino acid at every position was compared between telaprevir-exposed and -unexposed samples. Statistical significance was defined as P < 0.05 in Fisher's exact test; Bonferroni correction was used when multiple amino acid values or positions were explored simultaneously. For G3 analyses, unexposed samples comprised 102 G3a HCV sequences from combined public Los Alamos and internal HCV sequence databases. No formal statistical analyses were performed for G2, due to the low number of available sequences from unexposed samples.

G2 HCV-infected Patients

Antiviral Activity. During the investigational phase, vBT was observed in 6/9 G2 patients (66.7%) in the telaprevir monotherapy arm, compared with no vBT in the triple therapy and control arms (). One patient experienced vBT during the standard-treatment phase (control arm). One patient in the telaprevir monotherapy arm relapsed. HCV RNA profiles during telaprevir monotherapy are shown in Fig. 1a.

Table 1. Telaprevir antiviral activity in patients infected with either G2 or G3 HCV. Data are shown as n (%)

Baseline Virologic Data. Baseline population-based sequences were available for 22/23 G2 HCV-infected patients; no patient had mutations associated with decreased telaprevir susceptibility in G1 HCV. Clonal sequence analysis was conducted in the six patients with vBT: 1/27 clones in one patient had the T54A or R155K mutation at baseline; 1/22 clones in another patient had the A156V mutation at baseline (Fig. 2a).

Figure 2.

Variants detected by clonal sequencing in patients with vBT. Clonal sequence data are shown for (a) G2 and (b) G3 HCV-infected patients with vBT in the telaprevir monotherapy arm (20–40 clones/timepoint). Mutations at positions 36, 54, 155 and 156 were considered for this analysis. G, genotype

The mean G2 HCV telaprevir FC at baseline (vs wild type) was 2.4 (range, 0.2–6.6). For the seven telaprevir monotherapy patients with available phenotypic data, there was no clear relationship between baseline telaprevir FC and the change in HCV RNA from baseline to day 3 (Fig. 3a). However, the two patients with the lowest telaprevir FC at baseline had the greatest change in HCV RNA at day 3.

Genotypic Characterization of Viral Variants. Figure 4 shows HCV RNA, genotypic and phenotypic profiles over time for the six G2 HCV-infected patients (two G2b, one G2a and three G2 unknown subtype) in the telaprevir monotherapy arm with vBT.

In all five patients with vBT for whom sequences were available, emerging mutations associated with reduced telaprevir susceptibility in G1 HCV were observed using population sequencing. No sequence could be obtained for the day 15 sample of patient 5 due to low HCV RNA levels (101 IU/mL). At the time of vBT, T54A alone (n = 2), R155K alone (n = 1), A156S alone (n = 1) and the T54A+R155K combination (n = 1) were found. T54A was detected in one patient (patient 3) before vBT and A156S was detected in another patient (patient 1) after vBT. Clonal sequence analysis confirmed that in patients with vBT, mutations associated with reduced telaprevir susceptibility in G1 HCV emerged and became the dominant quasispecies (Fig. 2a).

In addition to the mutations associated with reduced susceptibility to telaprevir in G1 HCV, a number of other amino acid changes from baseline were identified in telaprevir monotherapy patients with vBT: Q32H, S146P, V163I, S166A and T179A (Fig. 4). However, no mutation appeared consistently across patients.

Of the six patients with vBT in the telaprevir monotherapy arm, five had undetectable HCV RNA at the end of peginterferon/ribavirin treatment, showing that the emerging variants remained sensitive to peginterferon/ribavirin. Three of these patients, including two with emerging mutations, achieved an SVR. For the patient who relapsed, no mutations associated with reduced susceptibility to telaprevir or other HCV protease inhibitors were detected at follow-up (week 24).[15]

Phenotypic Characterization of Viral Variants. Paired genotypic and phenotypic data were available for 3/6 G2 patients with vBT. At the time of vBT, two patients (patients 3 and 4, both G2b) had an emerging R155K mutation associated with a >9-fold increase from baseline in telaprevir FC. The same two patients had an emerging T54A mutation. In patient 3, T54A was not associated with an increase in telaprevir FC; while in patient 4, it was not present in the expression construct used for phenotyping. At the time of vBT, patient 6 (G2) had an emerging A156S mutation associated with a >48-fold increase from baseline in FC.

Introducing the R155K mutation into the baseline NS3 sequence of patients 3 and 4 by site-directed mutagenesis increased the telaprevir FC from 6.4 and 5.4 (without R155K) to >76.9 and >100, respectively. Introducing T54A into the baseline NS3 sequence of patient 4 increased the FC from 5.4 to 39.5. Introducing the A156S mutation into the baseline NS3 sequence of patients 1 (G2a) and 6 increased the FC from 1.2–1.6 to >58.8–>71.4.

G3 HCV-infected Patients

Antiviral Activity. During the investigational phase, vBT was observed in 3/8 G3 patients (37.5%) in the telaprevir monotherapy arm, compared with no vBT in the triple therapy and control arms (). As telaprevir monotherapy only produced slight decreases in HCV RNA levels (Fig. 1b), most patients did not meet the definition for vBT. Relapse was observed in two patients in the telaprevir monotherapy arm, three patients in the triple therapy arm and two control patients.

Table 1. Telaprevir antiviral activity in patients infected with either G2 or G3 HCV. Data are shown as n (%)

Baseline Virologic Data. Baseline population-based sequences were available for 24/26 G3 HCV-infected patients; no patient had mutations associated with decreased telaprevir susceptibility in G1 studies. No patient with vBT had known baseline mutations by clonal sequence analysis (Fig. 2b).

The mean telaprevir FC at baseline for G3 HCV (vs wild type) was 7.0 (range, 2.5–14.5). Unexpectedly, patients with the highest baseline telaprevir FC tended to show a greater change in HCV RNA between baseline and day 3 (Fig. 3b). However, patient numbers were small.

Genotypic Characterization of Viral Variants. All three telaprevir monotherapy patients with vBT had HCV subtype 3a (Fig. 5); R155K was present at vBT in two patients. In the third patient, no mutations associated with reduced telaprevir susceptibility in G1 studies were detected at vBT, but R155K was detected later. Clonal sequence analysis confirmed the results obtained by population sequencing (Fig. 2b). Two of the three patients with vBT had undetectable HCV RNA at the end of peginterferon/ribavirin treatment; one subsequently achieved an SVR, the other relapsed.

For patients without vBT, sequences of samples collected during the investigational phase were available for 4/5 patients in the telaprevir monotherapy arm and 1/7 patients in the triple therapy arm. No mutations associated with decreased telaprevir susceptibility in G1 HCV were detected by population sequencing. For patients who relapsed, sequences of samples were available for 4/5 patients in the telaprevir monotherapy and triple therapy arms. In patient 9, the R155K mutation was detected at time of relapse.

In addition to mutations associated with reduced susceptibility to telaprevir in G1, several other amino acid changes from baseline (T47A/T, A67A/V, K92K/N, A98A/T, S101A/T and P146S) were identified in G3 telaprevir monotherapy patients during or at the end of the investigational phase or at time of relapse. None of the NS3 region amino acid changes associated with telaprevir exposure reached statistical significance by Fisher's exact test.

Phenotypic Characterization of Viral Variants. Paired genotypic and phenotypic data were available for the three G3 patients with vBT. The emergence of R155K was associated with a >5-fold increase from baseline in FC.

Introduction of the R155K mutation into the NS3 baseline sequence of patients 7 and 9 (both G3a) by site-directed mutagenesis led to an FC increase from 9.4-10 (without R155K) to >58.8–>90.9 (with R155K).

Discussion

In treatment-naïve patients, telaprevir monotherapy had significant antiviral activity against G2 HCV but little or no activity against G3 HCV.[14] All vBTs occurred during telaprevir monotherapy, and most NS3 mutations emerging during breakthrough occurred at amino acid positions previously reported to be associated with reduced telaprevir susceptibility in G1 HCV. The finding that telaprevir monotherapy is associated with a high rate of vBT in G2/G3 HCV is similar to previous observations in G1 HCV,[15,16] and highlights the importance of combining telaprevir with peginterferon/ribavirin to control the emergence of resistant variants that remain susceptible to these agents.

In the telaprevir monotherapy arms, vBT was reported in six of nine patients with G2 HCV and three of eight patients with G3 HCV. Seven of these nine patients with vBT had undetectable HCV RNA at the end of subsequent treatment with peginterferon/ribavirin, and four achieved an SVR. The fact that G2 and G3 HCV variants with reduced telaprevir susceptibility remained sensitive to, and can be eradicated by, peginterferon/ribavirin extends similar observations with G1 HCV.[15]

Three mutations associated with reduced telaprevir susceptibility in G1 HCV (R155K, T54A, A156S) were seen in G2 HCV patients with vBT; two (R155K and T54A) were seen in G3 HCV patients with vBT. As in G1a HCV,[16] all emergent mutations required only a single nucleotide change from baseline. Several additional mutations were identified by the present analyses, but these did not occur consistently across patients and were not previously reported for other HCV protease inhibitors.[2,15,17] Moreover, none of these additional mutations occurred with significantly higher frequency than in HCV samples from patients not exposed to telaprevir. However, small sample sizes and a restricted diversity of subtypes limited the robustness of our findings and, for G2, statistical analyses could not be performed. Furthermore, in samples with these additional mutations, resistance could be explained by known telaprevir-resistant mutations using site-directed mutagenesis, although we had not formally introduced the additional mutations into baseline samples to determine their impact on telaprevir susceptibility. In spite of this, our data suggest a lack of G2- or G3-specific mutations for telaprevir resistance. Therefore, the resistance profile appears consistent across G1, G2 and G3.

The NS3 region of HCV shows some differences between genotypes. For example, position 36 of the NS3 region, identified as a site for telaprevir-resistant G1 mutations, contains leucine (L) in patients with G2 and G3 HCV, but valine (V) in G1 HCV. The impact of this amino acid change, if any, on telaprevir susceptibility is not fully understood and may be genotype and subtype dependent. For example, some G2 NS3 proteins carrying an 'L' at position 36 exhibited telaprevir FC values similar to those observed with G1 proteins carrying a 'V' at position 36 (Janssen Infectious Diseases BVBA, data on file). G3 NS3 proteins carrying an 'L' at position 36 exhibited on average higher FC values, although changes at other amino acid positions could have caused this decrease in susceptibility. This type of finding may underlie the differences observed in this study between G2 and G3 in baseline telaprevir FC, as well as the differences in viral activity of telaprevir against the two genotypes.

G2-infected patients with vBT during telaprevir therapy had subtype 2b, 2a or an unknown subtype, but none had 2c. Although patient numbers were small, this suggests that vBT may be less common with subtype 2c. All G3-infected patients with vBT had subtype 3a. The lack of vBT among subtype 3b patients could relate to the lack of telaprevir activity. The R155K mutation only emerged in G2-infected patients with subtype 2b; in G2a viruses, the emergence of a 'K' at position 155 would require a double nucleotide change. The R155K mutation was dominant in G3a HCV patients with vBT.

For patients with G2 HCV in the telaprevir monotherapy arm, the greatest HCV RNA decline from baseline to day 3 was observed in those with the highest telaprevir susceptibility at baseline. Conversely, in G3 patients, the greatest HCV RNA decline from baseline to day 3 was in those with the lowest baseline telaprevir susceptibility. Patient numbers were small, and the phenotype data were obtained with a biochemical assay, which is less reliable than a cell-based assay. However, this inverse correlation might relate to the low efficacy of telaprevir monotherapy in G3 patients. As previously observed in G1 HCV, emergence of the R155K mutation at vBT was associated with a high FC increase, and site-directed mutagenesis confirmed that R155K is associated with reduced susceptibility to telaprevir in G2/G3 patients. In addition, the A156S mutation was associated with reduced telaprevir susceptibility in G2 HCV.

Clonal sequence analysis confirmed that in most instances of vBT in patients with G2/G3 HCV, mutations associated with reduced telaprevir susceptibility in G1 HCV emerged and became the dominant quasispecies. In two patients, a mutation associated with reduced telaprevir susceptibility was detected at baseline in one clone. However, a mutation found in one clone of 22–27 clones can be the result of a PCR amplification error and not a true mutation present in the clonal population.

In addition to the limitations already mentioned for this study, the sample size was small, and a proportion of patients were unfortunately lost to follow-up. Small samples, however, are a common characteristic of proof-of-principle studies exploring the intrinsic activity of an antiviral agent.

In summary, telaprevir showed activity against G2 HCV in treatment-naïve patients, but limited or no activity against G3 HCV. The resistance profile of telaprevir in G2 and G3 HCV appears to involve similar amino acid substitutions as previously observed in G1 HCV. All cases of vBT occurred in the telaprevir monotherapy arms, highlighting the importance of combining telaprevir with peginterferon/ribavirin to avoid the emergence of resistant variants. A high proportion of patients who experienced vBT with telaprevir monotherapy still achieved an SVR after peginterferon/ribavirin treatment, indicating that the emergence of variants with reduced susceptibility to telaprevir can be controlled with these agents.

AcknowledgementsWe express our gratitude to the patients who participated in the study, as well as the study centre staff and Janssen Infectious Diseases personnel. We acknowledge Tom Westgate and Joanne Williams (Medical Writers, Gardiner-Caldwell Communications, Macclesfield, UK) for assistance in drafting the manuscript and collating author contributions. The clinical trial was sponsored by Janssen Pharmaceuticals and Vertex Pharmaceuticals.

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